Chronic obstructive pulmonary disease (COPD) is a major health problem worldwide and is increasing with a high prevalence, high morbidity and high cost. COPD is an important medical area for which biomarkers are needed for the study of the pathogenesis, the diagnosis and the clinical management. Exhaled breath condensate (EBC) can be collected with non-invasive methods and is a promising medium to develop biomarkers of COPD. The present research project is aimed at applying the most sensitive, selective and specific reference analytical techniques to the study of the composition of EBC in COPD patients. EBC levels of toxic metals and essential trace elements will be measured in COPD patients by electro-thermal atomic absorption spectroscopy (ETAAS) and inductively coupled plasma - mass spectrometry (ICP-MS). Such a novel approach (never done before) will be applied to identify and validate biomarkers of exposure and susceptibility to toxic metals, known to be contained in tobacco smoke, and probably playing a fundamental etiologic role in the pathogenetic path leading to COPD. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) will be used as a complementary approach to develop biomarkers of effect (aldehydes from lipid peroxidation) suitable for the long-term monitoring of COPD patients. This novel approach represents a significant advance over the analysis of alternative media (BALF, blood, serum, urine, hair), which are not as reliable (owing to interfering substances in the complex matrix) and reflect systemic rather than lung (target tissue) levels of both toxic metals and essential trace elements. Tobacco smoke and polluted environments substantially increase the lung burden of pneumotoxic chemicals, particularly heavy metals. Biomarkers of exposure (EBC levels of AI, Cd, Cr, Ni, Pb) should provide a quantitative estimate of the target tissue dose, thus distinguishing exposed from non-exposed controls. The lung response to inhaled pneumotoxic substances shows a high inter-individual variability as a function of different detoxifying capacity. Biomarkers of susceptibility will be developed relying on the consideration that some trace elements (Mn, Cu, Zn, Se, and Mo) are components of metalloproteins (superoxide dismutase, glutathione peroxidase, xanthine oxidase) known to modulate the response to toxic substances, possibly accounting for the limited proportion (15-20%) of smokers developing COPD. Biomarkers of susceptibility could be useful to identify and counsel people who are at increased risk of disease when exposed to tobacco smoke or environmental pollutants. Biomarkers of effect will be developed starting from aldehydes released into the EBC after lipid peroxidation in cell membranes. The pattern of aldehydes could distinguish membrane damage due to either oxidative stress or other mechanisms, such as a direct attack by free radicals. In summary, we propose a novel approach to the development of COPD biomarkers, relying on (i) noninvasive collection procedures to obtain (ii) a simple, interference-free matrix (EBC is practically water) to be analyzed by (iii) reference analytical methods, which are highly selective and specific (mainly, if not entirely, based on mass spectrometry), to determine (iv) stable analytes (heavy metals and trace elements). Through EBC biomarkers we sought to address two specific aims: a) to determine the validity of metals and aldehydes levels in EBC as biomarkers of exposure, of susceptibility and of effect in patients with COPD; b) to determine the clinical utility of EBC biomarkers as prognostic tools to predict the natural history of COPD. Establishing heavy and toxic metal levels in EBC as novel biomarkers of exposure, effect and susceptibility in patients with COPD could assist health care providers in devising new primary and secondary interventions, including drugs, to improve the quality of life and outcome of patients with COPD worldwide.